One of the goals of plant genetic engineering is to produce plants with agronomically important characteristics or traits. Recent advances in genetic engineering have provided the requisite tools to transform plants to contain and express foreign genes (Kahl et al. (1995) World Journal of Microbiology and Biotechnology 11:449-460). The technological advances in plant transformation and regeneration have enabled researchers to incorporate heterologous DNA into a plant's genome where it can then be expressed in the plant cell to exhibit the added characteristic(s) or trait(s). In one approach, expression of a novel gene that is not normally expressed in a particular plant or plant tissue may confer a desired phenotypic effect, in another approach, transcription of a gene, or part of a gene could be used to suppress transcription or translation of an endogenous gene. As a further example, a gene normally expressed in a particular tissue(s) or in a particular environmental or other condition, is expressed at higher levels than normal in the tissue, or under the condition.
In order to produce such a transgenic plant, a vector that includes a heterologous DNA polynucleotide that confers a phenotype when expressed in the plant is introduced into the plant cell. The vector also includes a promoter that functions in plants, operably linked to the DNA polynucleotide, often a promoter, not normally associated with the polynucleotide. The transformed plant cell is regenerated into a transgenic plant wherein the promoter controls expression of the introduced DNA polynucleotide to which the promoter is operably linked, and thus the DNA polynucleotide confers a change in some characteristic(s) of the plant. Isolated plant promoters are, therefore, useful tools for modifying plants to have desired phenotypic characteristics.
Because the promoter is a regulatory element that plays an integral part in the expression of a DNA polynucleotide, it would be advantageous to have a variety of promoters capable of selectively directing gene expression such that a DNA polynucleotide is transcribed efficiently at the selected time during plant growth and development, in the plant or portion thereof, and in the amount necessary to produce an effect. In one case, for example, it may be beneficial to have a gene product produced at a certain developmental stage, or in response to certain environmental or chemical stimuli.
A variety of different types or classes of promoters can be used for plant genetic engineering. Promoters can be classified on the basis of range or tissue specificity. For example, promoters referred to as constitutive promoters are capable of transcribing operably linked DNA polynucleotides efficiently and expressing the DNA polynucleotides in multiple tissues. Tissue-enhanced or tissue-specific promoters are operably linked to DNA polynucleotides normally transcribed in higher levels in certain plant tissues or specifically in certain plant tissues.
Relevant to this invention are promoters that provide enhanced expression in the abscission zone, apical meristem, roots, pod wall, and leaves of a plant. The abscission zone is a region of a plant wherein the production of hydrolytic enzymes lead to cell separation (Clements, American Journal of Botany 88:31 (2001)), which in turn leads to the shedding of leaves, flower, fruits and/or other plant parts. The process of abscission is well described in a review by Jane Taylor and Catherine Whitelaw (New Phytologist 151:323, 2001); “Abscission is the term used to describe the process of natural separation of organs from the parent plant. This may be part of the highly programmed development of a plant, or in response to environmental stress. It enables temperate plants to over-winter and hence survive, but in agricultural or horticultural environments premature abscission can lead to significant crop losses.” Promoters used to express genes that affect this process could be useful in modulating the loss of leaves, flowers, fruits and/or other plant parts and could lead to altered characteristics of the plant. In addition, the expression of genes in the abscission zone(s), and/or other key developmental organs and/or signaling pathways, provides scientists the ability to test approaches for increasing yield, abiotic and biotic stress tolerance, disease resistance, herbicide tolerance and resistance, and myriad other traits of agronomic importance.
Also of relevance are promoters that drive expression of genes in the apical meristem, pod wall, root, leaves, and/or flower of plants. Expression of DNA polynucleotides in these tissues could be useful to create plants with improved agronomic characteristics, including but not limited to increased yield.
By identifying and isolating a variety of plant promoters with desirable and/or unique gene expression patterns, it will be possible to operably link these promoters to DNA polynucleotides that provide advantageous agronomic effects in transgenic plants and plant cells.
Breeding permits the introduction of multiple traits into a single crop plant, also known as a gene stacking approach. In this approach, multiple genes conferring different characteristics of interest are introduced into a plant. It may be desirable when introducing multiple genes into a plant that each gene is modulated or controlled for the desired expression and that the promoter, and other regulatory elements, are diverse, to reduce the potential of gene silencing. Gene stacking can also be achieved through the introduction of a single covalently linked DNA polynucleotide that contains more than one promoter operably linked to more than one gene of interest, or with other promoter:gene combinations at different tDNA insertion sites, into a plant through transformation. It is contemplated that a plant comprising a recombinant DNA molecule of the present invention may be crossed, or stacked, with other traits such as herbicide resistance.